LAMP-2/CD107b Antibody (H4B4) - Azide and BSA Free

Western Blot: LAMP-2/CD107b Antibody (H4B4) - Azide and BSA Free [NBP2-80825] - Representative pictures of effect of I/R on DNA damage, cell survival, and inflammation. Western blot analysis expression level of Cathepsin B (*P < 0.05) in total protein extract from RVA of I/R treated vs. control group. Image collected and cropped by CiteAb from the following publication (//www.frontiersin.org/article/10.3389/fphar.2018.01034/full), licensed under a CC-BY license. Image from the standard format of this antibody.

Immunocytochemistry/Immunofluorescence: LAMP-2/CD107b Antibody (H4B4) - Azide and BSA Free [NBP2-80825] - T98G glioblastoma cells grown on #1.5 coverglass stained with a conjugated LAMP-2/CD107b (H4B4)AF647 antibody, counterstained with DAPI. Cathepsin D (green) staining also visible. This image was submitted via customer Review. Image using the Alexa Fluor 647 form of this antibody.

Immunohistochemistry: LAMP-2/CD107b Antibody (H4B4) - Azide and BSA Free [NBP2-80825] - Staining of LAMP-2/CD107b in human liver using DAB with hematoxylin counterstain. Image from the standard format of this antibody.

Flow Cytometry: LAMP-2/CD107b Antibody (H4B4) - Azide and BSA Free [NBP2-80825] - Intracellular flow cytometric staining of 1 x 10^6 HEK-293 cells using anti-LAMP-2/CD107b (dark blue). Isotype control shown in orange. An antibody concentration of 1 ug/1x10^6 cells was used. Image from the standard format of this antibody.

Western Blot: LAMP-2/CD107b Antibody (H4B4) - Azide and BSA Free [NBP2-80825] - Analysis of LAMP-2/CD107b expression in Whole Cell Lysates: 2) HeLa, 3) Jurkat, 4) U937, 5) HepG2, Tissue Extracts: 6) Human kidney, 7) Human lung, 8) Human liver, and 9) Human placenta. Image from the standard format of this antibody.

Western Blot: LAMP-2/CD107b Antibody (H4B4) - Azide and BSA Free [NBP2-80825] - Localization of TMEM135 to the mitochondria. The mitochondrial fraction isolated from the WT mouse brain show TMEM135 signals by immunoblotting. Following proteins were used as organelle markers: MFN2-mitochondria; LAMP2-lysosome; Lamin B1- nucleus; PDI- endoplasmic reticulum (ER). Image collected and cropped by CiteAb from the following publication (//elifesciences.org/articles/19264), licensed under a CC-BY license. Image from the standard format of this antibody.

Immunocytochemistry/Immunofluorescence: LAMP-2/CD107b Antibody (H4B4) - Azide and BSA Free [NBP2-80825] - HeLa cells were fixed for 10 minutes using 10% formalin and then permeabilized for 5 minutes using 1X TBS + 0.5% Triton X-100. The cells were incubated with anti-LAMP-2/CD107b [NBP2-22217] at a 1:100 dilution overnight at 4C and detected with an anti-mouse Dylight 488 (Green) at a 1:500 dilution. Actin was detected with Phalloidin 568 (Red) at a 1:200 dilution. Nuclei were counterstained with DAPI (Blue). Cells were imaged using a 40X objective. Image from the standard format of this antibody.

Flow Cytometry: LAMP-2/CD107b Antibody (H4B4) - Azide and BSA Free [NBP2-80825] - An intracellular stain was performed on U937 cells with HGF LAMP-2/CD107b [H4B4] Antibody NBP2-22217AF647 (blue) and a matched isotype control (orange). Cells were fixed with 4% PFA and then permeabilized with 0.1% saponin. Cells were incubated in an antibody dilution of 2.5 ug/mL for 30 minutes at room temperature. Both antibodies were directly conjugated to Alexa Fluor 647.

ELISA: LAMP-2/CD107b Antibody (H4B4) - Azide and BSA Free [NBP2-80825] - Anti-LAMP-2/CD107b was used to assess lysates from human SVG-A cells via ELISA, by loading the indicated mass of total protein per well. Antibodies tested were diluted 1:250, and used in conjunction with an HRP-conjugated secondary antibody. Signal was detected by luminescence. Image from verified customer review. Image from the standard format of this antibody.

Simple Western: LAMP-2/CD107b Antibody (H4B4) - Azide and BSA Free [NBP2-80825] - Lane view shows a specific band for LAMP-2/CD107b in 1.0 mg/ml of HeLa lysate. This experiment was performed under reducing conditions using the 12-230 kDa separation system. Image from the standard format of this antibody.

Western Blot: LAMP-2/CD107b Antibody (H4B4) - Azide and BSA Free [NBP2-80825] - Lysates of HeLa human cervical epithelial carcinoma parental cell line and LAMP2 knockout (KO) HeLa cell line. PVDF membrane was probed with Mouse Anti-Human LAMP-2/CD107b (H4B4) Monoclonal Antibody (Catalog # NBP2-22217) followed by HRP-conjugated Anti-Rabbit IgG Secondary Antibody (Catalog #HAF008). Specific band was detected for LAMP-2/CD107b at approximately 100 kDa (as indicated) in the parental HeLa cell line, but is not detectable in the knockout HeLa cell line. This experiment was conducted under reducing conditions. Image from the standard format of this antibody.

• Reactivity: Hu, Mu, Ch, Rt(-)
• Applications: WB, Simple Western, ELISA, Flow, ICC/IF, IHC, CyTOF-ready, KO
• Clone: H4B4
• Clonality: Monoclonal
• Host: Mouse
• Conjugate: Unconjugated
• Format: Azide and BSA Free
• Concentration: 1 mg/ml

LAMP-2/CD107b Antibody (H4B4) - Azide and BSA Free Summary

• Additional Information: Clone H4B4 was used by HLDA to establish CD designation.
• Immunogen: LAMP-2/CD107b Antibody (H4B4) was made using a human adherent spleen cells.
• Localization: Cell membrane; Single-pass type I membrane protein. Endosome membrane; Single-pass type I membrane protein. Lysosome membrane; Single-pass type I membrane protein. Note: This protein shuttles between lysosomes, endosomes, and the plasma membrane.
• Isotype: IgG1 Kappa
• Clonality: Monoclonal
• Host: Mouse
• Gene: LAMP2
• Purity: Protein G purified

Applications/Dilutions

• Dilutions:
  • CyTOF-ready
  • ELISA
  • Flow Cytometry 1 ug per million cells
  • Immunocytochemistry/ Immunofluorescence 1:10-1:100
  • Immunohistochemistry 1:100 - 1:200
  • Immunohistochemistry-Frozen 1:100 - 1:200
  • Immunohistochemistry-Paraffin 1:100 - 1:200
  • Knockout Validated
  • Simple Western 1:25
  • Western Blot 1:1000
• Application Notes: In Western blot, bands may be seen at ~40 kDa and 45 kDa representing the unglycosylated isoforms of LAMP2 and ~110 kDa representing the glycosylated form.
  
  In Simple Western only 10 - 15 ul of the recommended dilution is used per data point.
  See Simple Western Antibody Database for Simple Western validation: antibody dilution of 1:25. Separated by Size-Wes, Sally Sue/Peggy Sue. This antibody is CyTOF ready.
• Theoretical MW: 45 kDa.
  Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.

Reactivity Notes

Mouse reactivity reported in scientific literature (PMID: 27863209). Chicken reactivity reported in scientific literature (PMID: 30298003).

Packaging, Storage & Formulations

• Storage: Store at 4C short term. Aliquot and store at -20C long term. Avoid freeze-thaw cycles.
• Buffer: PBS
• Preservative: No Preservative
• Concentration: 1 mg/ml
• Purity: Protein G purified

Alternate Names for LAMP-2/CD107b Antibody (H4B4) - Azide and BSA Free

• CD107 antigen-like family member B
• CD107b antigen
• CD107b
• LAMP2
• LAMP-2
• LAMP2A
• LAMP-2A
• LAMPB
• LGP110
• LGP-96
• Lysosomal Associated Membrane Protein 2
• lysosomal-associated membrane protein 2
• lysosome-associated membrane glycoprotein 2
• Lysosome-associated membrane protein 2

Background

LAMP-2 (Lysosome-associated membrane protein 2) is a single-pass type I membrane protein that belongs to a family of membrane glycoproteins (~40 KDa). LAMP-2 protein is encoded by nine exons, with the first 8 exons and a portion of exon 9 encoding the highly glycosylated protein domains within the lysosomal lumen. The transmembrane and cytosolic carboxy-terminal domains of LAMP-2 are encoded by the remaining sequence of exon 9 and conform the receptor for targeting proteins to the lysosome. Splicing of exon 9 in the LAMP-2 pre mRNA leads to various splice forms with distinct cytosolic domains. Three splice variants, LAMP-2A, -2B and -2C, have been identified which shuttle between the plasma membrane, endosomal compartment and lysosomes (1). Tissue specific expression has been described for each LAMP-2 splice variant, with LAMP-2A being more ubiquitously expressed (e.g., placenta, lung, liver, pancreas and prostate), LAMP-2B predominantly expressed in skeletal muscle and LAMP-2C in brain tissue (1). All LAMP-2 splice variants participate in lysosomal degradation processes. LAMP-2A is the only variant that serves as a receptor targeting proteins for lysosomal degradation in chaperone-mediated autophagy (2,3). LAMP-2B is essential for macroautophagy in cardiomyocytes, where it facilitates autophagosome-lysosome fusion. LAMP-2B mutations underscore the myopathy and severe hypertrophic cardiomyopathy in Danon disease which results from deficits in autophagy (1, 4). Vasculopathy of coronary and cerebral arteries is a rare phenotype in Danon patients that is also associated with deficient autophagy processing of proteins and cellular organelles (5). LAMP2C serves as a receptor for DNA and RNA, facilitating their lysosomal degradation through DNA-autophagy and RNA-autophagy, respectively (1).

References

1. Rowland, T. J., Sweet, M. E., Mestroni, L., & Taylor, M. R. G. (2016). Danon disease - dysregulation of autophagy in a multisystem disorder with cardiomyopathy. Journal of Cell Science. https://doi.org/10.1242/jcs.184770

2. Alfaro, I. E., Albornoz, A., Molina, A., Moreno, J., Cordero, K., Criollo, A., & Budini, M. (2019). Chaperone mediated autophagy in the crosstalk of neurodegenerative diseases and metabolic disorders. Frontiers in Endocrinology. https://doi.org/10.3389/fendo.2018.00778

3. Schneider, J. L., & Cuervo, A. M. (2014). Autophagy and human disease: Emerging themes. Current Opinion in Genetics and Development. https://doi.org/10.1016/j.gde.2014.04.003

4. Chi, C., Leonard, A., Knight, W. E., Beussman, K. M., Zhao, Y., Cao, Y., Song, K. (2019). LAMP-2B regulates human cardiomyocyte function by mediating autophagosome lysosome fusion. Proceedings of the National Academy of Sciences of the United States of America. https://doi.org/10.1073/pnas.1808618116

5. Nguyen, H. T., Noguchi, S., Sugie, K., Matsuo, Y., Nguyen, C. T. H., Koito, H., Tsukaguchi, H. (2018). Small-Vessel Vasculopathy Due to Aberrant Autophagy in LAMP-2 Deficiency. Scientific Reports. https://doi.org/10.1038/s41598-018-21602-8

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Bioinformatics

• Gene Symbol: LAMP2